A major challenge in stem-cell mediated regenerative medicine is the development of defined culture systems for the maintenance of clinical-grade human embryonic stem (hES) cells. Using a feedback system control scheme, we identified a unique combination of three small molecule inhibitors that enables the maintenance of hES cells on a fibronectin-coated surface through single cell passaging. After 20 passages, the undifferentiated state of the hES cells was confirmed by OCT4, SSEA4 and NANOG expressions, while their pluripotent potential and genetic integrity were demonstrated by teratoma formation and normal karyotype, respectively. Our study attests to the power of feedback system control scheme in quickly pinpointing optimal conditions for desired biological activities and provides a chemically defined, scalable and single cell passaging culture system for hES cells.

Cell and microparticle separation in microfluidic systems has recently gained significant attention in sample preparations for biological and chemical studies. Microfluidic separation is typically achieved by applying differential forces on the target particles to guide them into different paths. This paper reviews basic concepts and novel designs of such microfluidic separators with emphasis on the use of non-inertial force fields, including dielectrophoretic force, optical gradient force, magnetic force, and acoustic primary radiation force. Comparisons of separation performances with discussions on physiological effects and instrumentation issues toward point-of-care devices are provided as references for choosing appropriate separation methods for various applications.

Microfluidic intracellular delivery approaches based on plasma membrane poration have shown promise for addressing the limitations of conventional cellular engineering techniques in a wide range of applications in biology and medicine. However, the inherent stochasticity of the poration process in many of these approaches often results in a trade-off between delivery efficiency and cellular viability, thus potentially limiting their utility. Herein, we present a novel microfluidic device concept that mitigates this trade-off by providing opportunity for deterministic mechanoporation (DMP) of cells en masse. This is achieved by the impingement of each cell upon a single needle-like penetrator during aspiration-based capture, followed by diffusive influx of exogenous cargo through the resulting membrane pore, once the cells are released by reversal of flow. Massive parallelization enables high throughput operation, while single-site poration allows for delivery of small and large-molecule cargos in difficult-to-transfect cells with efficiencies and viabilities that exceed both conventional and emerging transfection techniques. As such, DMP shows promise for advancing cellular engineering practice in general and engineered cell product manufacturing in particular.

Polydiacetylenes are a class of polymers with unique optical properties. Upon photopolymerization, monomers form a deep blue, nonfluorescent polymer, which transitions to a red, fluorescent polymer in response to various environmental factors such as pH, temperature, or molecular binding. The chromatic and emissive properties of polydiacetylenes have generated considerable popularity for their use in biosensing applications over the past three decades. The versatility of polydiacetylene forms has also allowed for a wide range of sensors including liposome bacterial sensors, films for detecting influenza virus, hydrogels for protein detection, and printed ink for the detection of volatile organic compounds. In this article, we review the wide range of techniques employed in the development of polydiacetylene sensors and summarize methods to modify, characterize, and analyze polydiacetylene-based sensing systems. Additionally, we discuss the recent directions of polydiacetylene materials outside of sensing applications as versatile tools in biomedicine and tissue engineering.

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We report a new polydiacetylene (PDA) sensor strip for simple visual detection of zinc ions in aqueous solution. The specificity of this sensor comes from Zn2+ DNA aptamer probes conjugated onto PDA. Effects of aptamer length and structure on the sensitivity of PDA’s color transition were first investigated. PDA conjugated with the optimal aptamer sequence was then coated onto a strip of polyvinylidene fluoride membrane and photopolymerized by UV exposure. The newly developed sensor successfully exhibited a blue-to-red chromatic change in a semi-quantitative manner in response to zinc ions. No discernable change was observed in solutions containing other common ions. Advantages of this sensor include its ease of fabrication, high specificity, and equipment-free detection, all of which are desirable for in-field applications and use in resource-limited settings.

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